Realizing the clinical utility of saliva for monitoring oral diseases.

In the era of personalized/precision health care, additional effort is being expended to understand the biology and molecular mechanisms of disease processes. How these mechanisms are affected by individual genetics, environmental exposures, and behavioral choices will encompass an expanding role in the future of optimally preventing and treating diseases. Considering saliva as an important biological fluid for analysis to inform oral disease detection/description continues to expand. This review provides an overview of saliva as a diagnostic fluid and the features of various biomarkers that have been reported. We emphasize the use of salivary biomarkers in periodontitis and transport the reader through extant literature, gaps in knowledge, and a structured approach toward validating and determine the utility of biomarkers in periodontitis. A summation of the findings support the likelihood that a panel of biomarkers including both host molecules and specific microorganisms will be required to most effectively identify risk for early transition to disease, ongoing disease activity, progression, and likelihood of response to standard periodontal therapy. The goals would be to develop predictive algorithms that serve as adjunctive diagnostic tools which provide the clinician and patient important information for making informed clinical decisions.

Transcriptomic features of programmed and inflammatory cell death in gingival tissues.

The local gingival tissue environment with homeostasis and tissue-destructive events of periodontitis demonstrates major changes in histological features and biology of the oral/sulcular epithelium, fibroblasts, vascular cells, inflammatory cell infiltration, and alveolar bone. OBJECTIVE: This study used an experimental periodontitis model to detail the gingival transcriptome related to cell death processes of pyroptosis, necroptosis, ferroptosis, and cuproptosis. MATERIALS AND METHODS: Healthy Macaca mulatta primates stratified by age, ≤3 years (young), 7-12 years (adolescent), 12-15 years (adult), and 17-23 years (aged), provided gingival tissue biopsies for microarray analysis focused on 257 genes representative of the four cell death processes and bacterial plaque samples for 16S rRNA gene analysis. RESULTS: Age differences in the profiles of gene expression in healthy tissues were noted for cuproptosis, ferroptosis, necroptosis, and pyroptosis. Major differences were then observed with disease initiation, progression, and resolution also related to the age of the animals. Distinct bacterial families/consortia of species were significantly related to the gene expression differences for the cell death pathways. CONCLUSIONS: These results emphasized age-associated differences in the gingival tissue molecular response to changes in the quality and quantity of bacteria accumulating with the disease process reflected in regulated cell death pathways that are both physiological and pathophysiological.

Gingival transcriptomic patterns of macrophage polarization during initiation, progression, and resolution of periodontitis.

Phenotypic and functional heterogeneity of macrophages is clearly a critical component of their effective functions in innate and adaptive immunity. This investigation hypothesized that altered profiles of gene expression in gingival tissues in health, disease, and resolution would reflect changes in macrophage phenotypes occurring in these tissues. The study used a nonhuman primate model to evaluate gene expression profiles as footprints of macrophage variation using a longitudinal experimental model of ligature-induced periodontitis in animals from 3 to 23 years of age to identify aging effects on the gingival environment. Significant differences were observed in distribution of expressed gene levels for M0, M1, and M2 macrophages in healthy tissues with the younger animals showing the least expression. M0 gene expression increased with disease in all but the aged group, while M1 was increased in adult and young animals, and M2 in all age groups, as early as disease initiation (within 0.5 months). Numerous histocompatibility genes were increased with disease, except in the aged samples. An array of cytokines/chemokines representing both M1 and M2 cells were increased with disease showing substantial increases with disease initiation (e.g. IL1A, CXCL8, CCL19, CCL2, CCL18), although the aged tissues showed a more limited magnitude of change across these macrophage genes. The analytics of macrophage genes at sites of gingival health, disease, and resolution demonstrated distinct profiles of host response interactions that may help model the disease mechanisms occurring with the formation of a periodontal lesion.

Biomarker panel discriminates diabetics with and without periodontitis pre- and post-therapy.

BACKGROUND AND OBJECTIVE: Biological regulators of periodontal inflammation, collagen degradation, and insulin resistance have not been determined in association with severity of periodontitis and response to periodontal treatment in diabetics. Our objective was to determine whether type 2 diabetes (T2DM) patients with periodontal disease present a distinct salivary biomarker profile compared with T2DM patients without periodontal disease and healthy subjects (without diabetes and periodontitis) pre- and post-nonsurgical therapy. METHODS: Clinical parameters of periodontal health and whole unstimulated saliva were collected from 92 participants (31 Not Periodontitis, NP; 32 T2DM without periodontitis, DWoP; and 29 with T2DM with periodontitis, DWP) at baseline. The T2DM groups received scaling and root planning (SRP) and provided saliva at 6-week follow-up. Salivary concentrations of interleukin (IL)-1ß, IL-6, matrix metalloproteinase-8 (MMP-8), and resistin were measured by immunoassay. RESULTS: The DWP group had significantly more disease and higher salivary concentrations at baseline for IL-1ß, MMP-8, and resistin (p's < .01) compared with DWoP and NP. SRP resulted in significant improvement in periodontal parameters for the T2DM groups; however, more disease persisted (p < .001), and IL-1ß, MMP-8, and resistin concentrations remained significantly higher in the DWP than the DWoP group (p < .01) at 6 weeks post-treatment. Principal component analysis demonstrated the DWoP group appeared more biologically similar to the NP group than the DWP group. Concentrations of these salivary biomarkers increased with increasing periodontal disease severity (p < .05) in this study population. CONCLUSION: Salivary concentrations of IL-1ß, MMP-8, and resistin appear to serve as biomarkers of periodontal status pre- and post-treatment, irrespective of diabetes status.

Dietary Strawberries Improve Serum Metabolites of Cardiometabolic Risks in Adults with Features of the Metabolic Syndrome in a Randomized Controlled Crossover Trial.

Dietary strawberries have been shown to improve cardiometabolic risks in multiple clinical trials. However, no studies have reported effects on serum metabolomic profiles that may identify the target pathways affected by strawberries as underlying mechanisms. We conducted a 14-week randomized, controlled crossover study in which participants with features of metabolic syndrome were assigned to one of the three arms for four weeks separated by a one-week washout period: control powder, 1 serving (low dose: 13 g strawberry powder/day), or 2.5 servings (high dose: 32 g strawberry powder/day). Blood samples, anthropometric measures, blood pressure, and dietary and physical activity data were collected at baseline and at the end of each four-week phase of intervention. Serum samples were analyzed for primary metabolites and complex lipids using different mass spectrometry methods. Mixed-model ANOVA was used to examine differences in the targeted metabolites between treatment phases, and LASSO logistic regression was used to examine differences in the untargeted metabolites at end of the strawberry intervention vs. the baseline. The findings revealed significant differences in the serum branched-chain amino acids valine and leucine following strawberry intervention (high dose) compared with the low-dose and control phases. Untargeted metabolomic profiles revealed several metabolites, including serum phosphate, benzoic acid, and hydroxyphenyl propionic acid, that represented improved energy-metabolism pathways, compliance measures, and microbial metabolism of strawberry polyphenols, respectively. Thus, dietary supplementation of strawberries significantly improves the serum metabolic profiles of cardiometabolic risks in adults.

Streptococcus gordonii-Induced miRNAs Regulate CCL20 Responses in Human Oral Epithelial Cells.

The mechanisms through which oral commensal bacteria mitigates uncontrolled inflammatory responses of the oral mucosa remain unknown. Here, we show that representative oral bacterial species normally associated with oral health [S. gordonii (Sg), V. parvula (Vp), A. naeslundii (An), C. sputigena (Cs), and N. mucosa (Nm)] enhanced differential chemokine responses in oral epithelial cells (OECs), with some bacteria (An, Vp, and Nm) inducing higher chemokine levels (CXCL1, CXCL8) than others (Sg, Cs). Although all bacterial species (except Cs) increased CCL20 mRNA levels consistent with protein elevations in cell lysates, only An, Vp, and Nm induced higher CCL20 secretion, similar to the effect of the oral pathogen F. nucleatum (Fn). In contrast, most CCL20 remained associated with OECs exposed to Sg and negligible amounts released into the cell supernatants. Consistently, Sg attenuated An-induced CCL20. MiR-4516 and miR-663a were identified as Sg-specifically induced miRNAs modulating validated targets of chemokine-associated pathways. Cell transfection with miR-4516 and miR-663a decreased An- and Fn-induced CCL20. MiRNA upregulation and attenuation of An-induced CCL20 by Sg were reversed by catalase. Up-regulation of both miRNAs was specifically enhanced by oral streptococci H2O2-producers. These findings suggest that CCL20 levels produced by OECs in response to bacterial challenge are regulated by Sg-induced miR-4516 and miR-663a in a mechanism that involves hydrogen peroxide. This type of molecular mechanism could partly explain the central role of specific oral streptococcal species in balancing inflammatory and antimicrobial responses given the critical role of CCL20 in innate (antimicrobial) and adaptive immunity (modulates Th17 responses).

Gingival tissue antibody gene utilization in aging and periodontitis.

OBJECTIVE: This study used a nonhuman primate model of ligature-induced periodontitis to document the characteristics of immunoglobulin (Ig) gene usage in gingival tissues with disease and affected by age. BACKGROUND: Adaptive immune responses to an array of oral bacteria are routinely detected in local gingival tissues and the systemic circulation across the human population. The level and diversity of antibody increases with periodontitis, reflecting the increased quantity of B cells and plasmacytes in the tissues at sites of periodontal lesions. METHODS: Macaca mulatta (n = 36) in four groups (young - ≤3 years; adolescent >3-7 years; adult - 12-15 years; aged - 17-23 years) were used in this study. Gingival tissues were sampled at baseline (health), 2 weeks (initiation), 1 and 3 months (progression), and 5 months (resolution) of the lesion development and transcriptomic analysis included 78 Ig-related genes. RESULTS: The results demonstrated extensive variation in Ig gene usage patterns and changes with the disease process that was substantially affected by the age of the animal. Of note was that the aged animals generally demonstrated elevated expression on multiple Ig genes even in the baseline/healthy gingival tissues. The expression levels revealed 5 aggregates of Ig gene change profiles across the age groups. The number of gene changes were greatly increased in adult animals with the initiation of disease, while the young and adolescent animals showed extensive changes with disease progression. Elevated Ig gene transcripts remained with disease resolution except in the aged animals. The response profiles demonstrated selective heavy/light change gene transcripts that differed with age and clustering of the transcript expression was dominated by the age of the animals. CONCLUSIONS: The results suggested potential critical variations in the molecular aspects of Ig gene expression in gingival tissues that can contribute to understanding the kinetics of periodontal lesions, as well as the variation in episodes, rapidity of progression, and role in resolution that are impacted by age.

Oral microbiome interactions with gingival gene expression patterns for apoptosis, autophagy and hypoxia pathways in progressing periodontitis.

Oral mucosal tissues must react with and respond to microbes comprising the oral microbiome ecology. This study examined the interaction of the microbiome with transcriptomic footprints of apoptosis, autophagy and hypoxia pathways during periodontitis. Adult Macaca mulatta (n = 18; 12-23 years of age) exhibiting a healthy periodontium at baseline were used to induce progressing periodontitis through ligature placement around premolar/molar teeth. Gingival tissue samples collected at baseline, 0·5, 1 and 3 months of disease and at 5 months for disease resolution were analysed via microarray. Bacterial samples were collected at identical sites to the host tissues and analysed using MiSeq. Significant changes in apoptosis and hypoxia gene expression occurred with initiation of disease, while autophagy gene changes generally emerged later in disease progression samples. These interlinked pathways contributing to cellular homeostasis showed significant correlations between altered gene expression profiles in apoptosis, autophagy and hypoxia with groups of genes correlated in different directions across health and disease samples. Bacterial complexes were identified that correlated significantly with profiles of host genes in health, disease and resolution for each pathway. These relationships were more robust in health and resolution samples, with less bacterial complex diversity during disease. Using these pathways as cellular responses to stress in the local periodontal environment, the data are consistent with the concept of dysbiosis at the functional genomics level. It appears that the same bacteria in a healthy microbiome may be interfacing with host cells differently than in a disease lesion site and contributing to the tissue destructive processes.

Dietary Blueberry and Soluble Fiber Supplementation Reduces Risk of Gestational Diabetes in Women with Obesity in a Randomized Controlled Trial.

BACKGROUND: Gestational diabetes mellitus (GDM) is a growing public health concern and maternal obesity and poor dietary intakes could be implicated. Dietary polyphenols and fiber mitigate the risk of diabetes and its complications, but little is known about their efficacy in preventing GDM. OBJECTIVES: We examined the effects of whole blueberry and soluble fiber supplementation on primary outcomes of cardiometabolic profiles in women at high risk of developing GDM. METHODS: Women (n = 34; mean ± SD age: 27 ± 5 y; BMI: 35.5 ± 4.0 kg/m2; previous history of GDM ∼56%; Hispanic ∼79%) were recruited in early pregnancy (<20 weeks of gestation) and randomly assigned to 1 of the following 2 groups for 18 wk: intervention (280 g whole blueberries and 12 g soluble fiber per day) and standard prenatal care (control). Both groups received nutrition education and maintained 24-h food recalls throughout the study. Data on anthropometrics, blood pressure, and blood samples for biochemical analyses were collected at baseline (<20 weeks), midpoint (24-28 weeks), and end (32-36 weeks) of gestation. Diagnosis of GDM was based on a 2-step glucose challenge test (GCT). Data were analyzed using a mixed-model ANOVA. RESULTS: Maternal weight gain was significantly lower in the dietary intervention than in the control group at the end of the trial (mean ± SD: 6.8 ± 3.2 kg compared with 12.0 ± 4.1 kg, P = 0.001). C-reactive protein was also lower in the intervention than in the control group (baseline: 6.1 ± 4.0 compared with 6.8 ± 7.2 mg/L; midpoint: 6.1 ± 3.7 compared with 7.5 ± 7.3 mg/L; end: 5.5 ± 2.2 compared with 9.5 ± 6.6 mg/L, respectively, P = 0.002). Blood glucose based on GCT was lower in the intervention than in the control (100 ± 33 mg/dL compared with 131 ± 40 mg/dL, P < 0.05). Conventional lipids (total, LDL, and HDL cholesterol and triglycerides) did not differ between groups over time. No differences were noted in infant birth weight. CONCLUSIONS: Whole blueberry and soluble fiber supplementation may prevent excess gestational weight gain and improve glycemic control and inflammation in women with obesity.This trial was registered at clinicaltrials.gov as NCT03467503.

Variations in IgG antibody subclass responses to oral bacteria: Effects of periodontal disease and modifying factors.

BACKGROUND AND OBJECTIVE: Local and systemic IgG antibodies or oral bacteria have been described with periodontitis. We extended these observations by assessing the impact of a range of intrinsic factors on serum IgG subclass antibodies to both commensal and pathogenic oral bacteria that would contribute to variations in immune protection or disease susceptibility in periodontitis have not been described. METHODS: Subjects (n = 278) were classified as healthy, gingivitis, or periodontitis and categorized as mild, moderate, and severe periodontitis. Demographic stratification included sex, age, race/ethnicity, smoking, and obesity. Whole formalin-fixed bacteria were used as antigens to detect serum immunoglobulin (Ig)G subclass antibody levels using an ELISA. RESULTS: The greatest differences in variations in IgG subclasses occurred in periodontitis versus health or gingivitis to bacteria considered oral pathogens (eg, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, and Treponema denticola) with IgG1, IgG2, and IgG4 increased by three- to sevenfold with Pg. Differences in subclass levels and distribution were also observed related to disease severity, particularly related to individual subclass responses to Pg. Examination of the overall population showed that females had elevated antibody, reflected by elevated IgG2 amounts/proportions. The older group of subjects demonstrated elevated antibody to multiple oral bacteria, lacking any particular subclass pattern. IgG2 antibody to Aa and Pg was increased in smokers. Multiple IgG subclass antibody levels to oral pathogens were significantly decreased in the obese subset within this population. CONCLUSION: This investigation identified patterns of IgG subclass antibody responses to oral bacteria and demonstrated substantial effects of disease impacting the level and subclass distribution of antibody to an array of oral bacteria. Altered subclass antibody profiles most often in IgG2 levels and for antibody to P. gingivalis were found related to sex, age, disease severity, race/ethnicity, smoking, and obesity to both pathogens and commensal bacteria.

Gingival tissue autophagy pathway gene expression profiles in periodontitis and aging.

OBJECTIVE: We hypothesized that autophagy-related genes will be differentially expressed in periodontitis, suggesting an impaired gingival autophagic response associated with disease. BACKGROUND: Autophagy is a cellular physiologic mechanism to maintain tissue homeostasis, while deficient autophagic responses increase inflammation and susceptibility to infection. METHODS: Rhesus monkeys [<3 years to 23 years of age (n = 34)] were examined for periodontal health and naturally occurring periodontitis. Gingival tissues samples were obtained from healthy or diseased sites, total RNA was isolated, and the Rhesus Gene Chip 1.0 ST (Affymetrix) was used for gene expression analysis of 150 autophagy-related genes. RESULTS: Comparison of expression levels with adult healthy tissues demonstrated a rather limited number of individual genes that were significantly different across the age-groups. In contrast, with periodontitis in the adults and aged animals, about 15% of the genes were significantly increased or decreased. The differences were reflected in the mTOR complex (5/12), ULK1/ATG1 complex (5/9), PI3K complex (5/21), ATG9 complex (2/7), ATG12 conjugation/LC3 lipidation (7/22), and lysosome fusion/vesicle degradation [LF/VD (5/10)] activities within the broader autophagic pathway. The genes most greatly altered in gingival tissues of naturally occurring periodontitis were identified in the ATG12 and LF/VD pathways that approximated 50% of the genes in each of those categories. While healthy gingival aging did not appear to reflect altered autophagy gene expression, substantial differences were noted with periodontitis irrespective of the age of the animals. Future studies into the role of autophagy in periodontitis and could offer potential new therapeutic strategies to prevent and/or treat periodontal disease.

Salivary biomarkers for discriminating periodontitis in the presence of diabetes.

AIM: Salivary biomarkers can help in assessment of periodontitis; however, concentrations may be altered in the presence of diabetes. Hence, the ability of salivary biomarkers to discriminate periodontally healthy type II diabetics (T2DM) from T2DM who have periodontitis was examined. METHODS: Ninety-two participants (29 with T2DM with chronic periodontitis, DWP; 32 T2DM without chronic periodontitis, DWoP; and 31 Not Periodontitis, NP) provided saliva and clinical parameters of periodontal health were recorded. Salivary concentrations of interleukin (IL)-1ß, IL-6, matrix metalloproteinase-8 (MMP-8), macrophage inflammatory protein-1α (MIP-1α), adiponectin and resistin were measured by immunoassay. RESULTS: Salivary analyte concentrations for IL-1ß, MMP-8 and resistin correlated with clinical parameters of periodontitis, with MMP-8 demonstrating the strongest positive correlation with PD ≥5 mm (p < 0.0001). Periodontal health was reflected in salivary analyte concentrations by group, with concentrations of IL-1ß and MMP-8 showing significant associations with periodontitis (p ≤ 0.04) that increased in concentration from health to DWoP to DWP. Odds ratio (OR) analyses showed that MMP-8 discriminated periodontitis from NP (OR of 8.12; 95% CI: 1.01-65.33; p = 0.03) and in the presence of T2DM (DWP vs DWoP, OR = 5.09; 95% CI: 1.24-20.92; p = 0.03). CONCLUSION: Salivary MMP-8 and IL-1ß discriminate periodontitis in T2DM.

Relationship between herpesviruses and periodontal disease progression.

AIM: To investigate the role of Epstein-Barr virus (EBV), cytomegalovirus (CMV), and anaerobic bacteria in the progression of periodontitis. METHODS: Eighty-one adults with generalized moderate to severe periodontitis were randomly assigned to: oral hygiene or scaling and root planning ± placebo or polyunsaturated fatty acids fish oil. Subgingival plaque samples collected from three healthy and three disease sites at weeks 0, 16, and 28 and from sites demonstrating disease progression were analysed for EBV, CMV, P. gingivalis (Pg), T. forsythia (Tf), and T. denticola (Td) DNA using quantitative polymerase chain reaction. RESULTS: Cytomegalovirus was detected in 0.3% (4/1454) sites. EBV was present in 12.2% of healthy sites (89/728) and 27.6% disease sites (201/726; p < .0001), but was in low copy number. Disease progression occurred in 28.4% of participants (23/81) and developed predominantly at sites identified as diseased (75/78; 96.2%). CMV and EBV were not associated with disease progression (p = .13) regardless of treatment. In contrast, disease sites were associated with higher levels of Pg, Td, Tf, and total bacteria, and sites that exhibited disease progression were associated with an abundance of Td and Tf (p < .04). CONCLUSION: Disease progression was associated with Gram-negative anaerobic bacteria; not EBV or CMV.

Periodontal disease susceptible matrilines in the Cayo Santiago Macaca mulatta macaques.

OBJECTIVE AND BACKGROUND: The expression of periodontitis, including age of onset, extent, and severity is considered to represent an interaction of the individual's oral microbiome and host response to the microbial challenge that is modified by both genetics and environmental factors. The aim of this study was to determine the distribution of periodontitis in a population of nonhuman primates, to document features of familial distribution that could reflect heritability and transmission of microbes with enhanced virulence. MATERIAL AND METHODS: This report presents our findings from evaluation of periodontal disease bone defects in skulls from 569 animals (5-31 years of age) derived from the skeletons of the rhesus monkeys (Macaca mulatta) of Cayo Santiago derived from eight matrilines over 6-9 generations. The distance from the base of alveolar bone to the cemento-enamel junction on 1st /2nd premolars and 1st /2nd molars from all four quadrants was evaluated as a measure of periodontal disease. Additionally, we documented the presence of periodontitis in 79 living descendants within these matrilines. RESULTS: The results demonstrated an increased extent and severity of periodontitis with aging across all matrilines. Extensive heterogeneity in disease expression was observed among the animals and this was linked to specific periodontitis susceptible matrilines. Moreover, we identified some matrilines in which the members appeared to show some resistance to more severe disease, even with aging. CONCLUSION: Linking these disease variations to multigenerational matriarchal family units supported familial susceptibility of periodontitis. This familial disease relationship was reinforced by the distribution of naturally-occurring periodontitis in the living descendants.

Biological response to peri-implantitis treatment.

OBJECTIVE: To investigate biological markers of peri-implantitis (PIP) in crevicular fluid before and after surgical and antimicrobial therapy. MATERIAL AND METHODS: Forty-eight participants (24 healthy implants and 24 PIP) were clinically evaluated, and peri-implant crevicular fluid (PICF) samples were collected at baseline for both groups, and at 3-months after surgical and antimicrobial treatment (ie, n = 21 PIP completers). Samples were analyzed for interleukin-1ß (IL-1ß), matrix metalloproteinase-8 (MMP-8), and macrophage inflammatory protein-1α (MIP-1α) using immunoassay and the results compared between groups. RESULTS: Peri-implantitis sites at baseline demonstrated significantly higher mean periodontal probing depths, percentage bleeding on probing (P ≤ 0.001), and mean IL-1ß concentration in PICF compared to healthy implant sites (17.9 vs 1.7 pg/µL; P = 0.02). Three months after treatment, periodontal probing depths, bleeding on probing, suppuration (P < 0.05), and the mean concentration of MMP-8 decreased significantly compared with baseline (12.1 vs 6.7 ng/µL, P = 0.04). MIP-1α concentrations showed no differences between the groups. CONCLUSION: Elevated concentrations of IL-1ß in PICF were consistent with PIP. A decrease in MMP-8 concentration in PICF at three months after treatment is consistent with a healing biological response.

Biologic modelling of periodontal disease progression.

AIM: To investigate the synergistic role of biologic markers from saliva, serum and plaque in modelling periodontitis disease progression. MATERIAL AND METHODS: This longitudinal study evaluated characteristics of disease progression in 114 patients with generalized moderate to severe periodontitis. The primary outcome was detection of sites with progressing attachment loss sites over 6 months in patients who received scaling and root planing or oral hygiene only. The predictive potential of 27 biomarkers in serum, whole saliva and subgingival plaque was evaluated using three classification algorithms (Support Vector Machines; Naïve Bayes Classifier; and Linear Discriminant Analysis) within an ensemble predictive modelling framework. RESULTS: Disease progression occurred in 24.6% of subjects (28/114). Predictive modelling using Naïve Bayes Classifier identified progressors best with sensitivity of ~89%. The use of the three classification algorithms revealed the concerted role of salivary matrix metalloproteinase-8, serum biomarkers (serum amyloid P, matrix metalloproteinase 1, bactericidal permeability-increasing protein, isoprostane) along with levels of Porphryomonas gingivalis and Tannerella forsythia in plaque in predicting progressors. CONCLUSIONS: Synergistic utility of baseline bacterial and inflammatory biomarkers from saliva, serum and plaque predicted disease progression.

Hypoxia-inducible transcription factors, HIF1A and HIF2A, increase in aging mucosal tissues.

Hypoxia (i.e. oxygen deprivation) activates the hypoxia-signalling pathway, primarily via hypoxia-inducible transcription factors (HIF) for numerous target genes, which mediate angiogenesis, metabolism and coagulation, among other processes to try to replenish tissues with blood and oxygen. Hypoxia signalling dysregulation also commonly occurs during chronic inflammation. We sampled gingival tissues from rhesus monkeys (Macaca mulatta; 3-25 years old) and total RNA was isolated for microarray analysis. HIF1A, HIF1B and HIF2A were significantly different in healthy aged tissues, and both HIF1A and HIF3A were positively correlated with aging. Beyond these transcription factor alterations, analysis of patterns of gene expression involved in hypoxic changes in tissues showed specific increases in metabolic pathway hypoxia-inducible genes, whereas angiogenesis pathway gene changes were more variable in healthy aging tissues across the animals. With periodontitis, aging tissues showed decreases in metabolic gene expression related to carbohydrate/lipid utilization (GBE1, PGAP1, TPI1), energy metabolism and cell cycle regulation (IER3, CCNG2, PER1), with up-regulation of transcription genes and cellular proliferation genes (FOS, EGR1, MET, JMJD6) that are hypoxia-inducible. The potential clinical implications of these results are related to the epidemiological findings of increased susceptibility and expression of periodontitis with aging. More specifically the findings describe that hypoxic stress may exist in aging gingival tissues before documentation of clinical changes of periodontitis and, so, may provide an explanatory molecular risk factor for an elevated capacity of the tissues to express destructive processes in response to changes in the microbial biofilms characteristic of a more pathogenic microbial challenge.

Environmental lead effects on gene expression in oral epithelial cells.

BACKGROUND AND OBJECTIVE: Host responses in periodontitis span a range of local and emigrating cell types and biomolecules. Accumulating evidence regarding the expression of this disease across the population suggests some component of genetic variation that controls onset and severity of disease, in concert with the qualitative and quantitative parameters of the oral microbiome at sites of disease. However, there remains little information regarding the capacity of accruing environmental stressors or modifiers over a lifespan at both the host genetic and microbial ecology levels to understand fully the population variation in disease. This study evaluated the impact of environmental lead exposure on the responses of oral epithelial cells to challenge with a model pathogenic oral biofilm. METHODS AND RESULTS: Using NanoString technology to quantify gene expression profiles of an array of 511 host response-associated genes in the epithelial cells, we identified an interesting primary panel of basal responses of the cells with numerous genes not previously considered as major response markers for epithelial cells, eg, interleukin (IL)-32, CTNNB1, CD59, MIF, CD44 and CD99. Even high levels of environment lead had little effect on these constitutive responses. Challenge of the cells with the biofilms (Streptococcus gordonii/Fusobacterium nucleatum/Porphyromonas gingivalis) resulted in significant increases in an array of host immune-related genes (134 of 511). The greatest magnitude in differential expression was observed with many genes not previously described as major response genes in epithelial cells, including IL-32, CD44, NFKBIA, CTSC, TNFAIP3, IL-1A, IL-1B, IL-8 and CCL20. The effects of environmental lead on responses to the biofilms were mixed, although levels of IL-8, CCL20 and CD70 were significantly decreased at lead concentrations of 1 and/or 5 µmol/L. CONCLUSION: The results provided new information on a portfolio of genes expressed by oral epithelial cells, targeted substantial increases in an array of immune-related genes post-biofilm challenge, and a focused impact of environmental lead on these induced responses.

Ageing effects on humoral immune responses in chronic periodontitis.

Periodontal disease is a dominant global bacterial infection that increases with ageing. AIM: This report focuses on host adaptive immune responses in periodontitis. While experimental models and humans diagnosed with periodontitis demonstrate an antigenic specificity for particular oral bacteria, we have a limited understanding of (i) how ageing affects the adaptive immune responses to these bacteria that chronically colonize the oral cavity for decades prior to disease expression and (ii) how the magnitude and specificity of the response interface with pathogens that emerge within the bacterial ecology during exacerbations of disease. MATERIALS AND METHODS: Serum antibody levels to a group of pathogenic and commensal oral bacteria were measured in a population of individuals from 21 to 74 years of age, stratified based on clinical status of the periodontium, smoking and sex. RESULTS: Clinical parameters were not significantly different within health, gingivitis or periodontitis groups related to age. Antibody to oral pathogens and commensals was similar in different age groups in each of the clinical categories, with no age correlation noted in the periodontitis patients. CONCLUSIONS: The adaptive immune responses to oral bacteria that chronically colonize the oral cavity appear generally unaffected by age, but clearly are linked to the extent of disease.

Odontogenic abscesses in rhesus macaques (Macaca mulatta) of Cayo Santiago.

OBJECTIVES: Odontogenic abscesses are one of the most common dental diseases causing maxillofacial skeletal lesions. They affect the individual's ability to maintain the dental structures necessary to obtain adequate nutrition for survival and reproduction. In this study, the prevalence and pattern of odontogenic abscesses in relation to age, sex, matriline, and living periods were investigated in adult rhesus macaque skeletons of the free-ranging colony on Cayo Santiago, Puerto Rico. MATERIALS AND METHODS: The skulls used for this study were from the skeletons of 752 adult rhesus macaques, aged 8-31 years, and born between 1951 and 2000. They came from 66 matrilines ranging from 1 to 88 individuals. Fistulae or skeletal lesions caused by odontogenic abscesses drainage, carious lesions, tooth fractures, tooth loss, and alveolar resorption were evaluated visually. RESULTS: Seventy-two specimens (9.57%) had odontogenic abscesses of varying severity. Males had a significantly higher prevalence than females. The prevalence of odontogenic abscesses in several matrilines was significantly higher than in the population as a whole. Animals born between 1950 and 1965 tended to have a higher prevalence of odontogenic abscesses than those born in later periods. DISCUSSION: These results suggest that oral pathologies, such as dental and periodontal abscesses in rhesus macaques are fairly common, which may indicate familial effects interwoven with ecological and social factors. The closeness of the rhesus and human genomes allows insights to understand of the epidemiology of these diseases in the human population. Further assessment of the role played by environmental and familial factors on rhesus oral health and disease are warranted.